Abstract
Urinary pseudouridine levels have been proposed as diagnostic biomarkers for various malignancies; however, their association with colorectal cancer (CRC) remains unclear. This study investigates the molecular mechanisms underlying pseudouridine-related genes (PRGs) in CRC. The study incorporated a training cohort (TCGA-CRC), a validation cohort (GSE87211), a single-cell dataset (GSE200997), and PRGs retrieved from public databases. Quality control was performed on the single-cell dataset, followed by cell type annotation. Differentially expressed genes (DEGs) across distinct cell populations were identified. Weighted gene co-expression network analysis (WGCNA) was employed to screen module genes strongly correlated with PRG scores. DEGs between tumor and normal samples in the training cohort were also determined. Candidate genes were selected by intersecting DEGs from key cell types, tumor-normal comparisons, and WGCNA-derived module genes. A prognostic risk model was constructed using Cox regression analyses. Independent prognostic factors were identified through univariate and multivariate Cox analyses, integrating clinical parameters and risk scores, to establish a prognostic nomogram. Comparative analyses of mutation profiles, immune infiltration, and functional pathways were conducted between high- and low-risk groups, and molecular mechanisms of prognostic genes were explored. Additionally, pseudo-temporal trajectory analysis was applied to assess prognostic gene expression dynamics in key cell types. Seven cell types were annotated in the single-cell dataset, with T cells and epithelial cells representing predominant and functionally significant populations. A total of 116 candidate genes were identified by overlapping 4,762 DEGs from T cells, 4,525 DEGs from epithelial cells, 9,772 tumor-normal DEGs, and 2,990 module genes. A prognostic risk model incorporating three PRGs—BCL10, TAF1B, and WWTR1—was developed and validated across training and validation cohorts. Risk score, age, T stage, N stage, and tumor stage were recognized as independent prognostic factors for constructing the nomogram. Pseudo-temporal trajectory analysis revealed that TAF1B expression was relatively elevated at the terminal differentiation phase in epithelial cells. A pseudouridine-related prognostic model based on three PRGs was established and validated, offering a potential reference for CRC treatment and risk stratification.
Data availability
The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found below: https://portal.gdc.cancer.gov, The Cancer Genome Atlas; https://www.ncbi.nlm.nih.gov/geo, The Gene Expression Omnibus; https://www.gsea-msigdb.org/gsea/msigdb/index.jsp, The Molecular Signatures Database.
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Funding
This project was supported by the National Natural Science Foundation of China (No. 82260543), Natural Science Foundation of Ningxia (2023AAC05058, 2024AAC03557), and Scientific Research Foundation of Fujian Provincial Hospital, China (No.2020YJ04).
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The research was planned by TJ. The data was analyzed and the original article was written by ZJW and LYM. JZC and STL were responsible for the in vitro experimental validation and contributed to data analysis and interpretation. RXM and JW gathered references and reviewed the paper. The data was gathered by HYL and ZXZ. The final manuscript was reviewed and approved by all authors.
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Written informed consent was obtained from each patient before surgery and all study protocols were approved by the Ethics Committee for Clinical Research of General Hospital of Ningxia Medical University (Reference Number : KYLL-2022-0800). All methods were carried out in accordance with relevant guidelines and regulations/Declaration of Helsinki.
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Wang, Z., Ma, L., Cao, J. et al. A pseudouridine-related prognostic model of colorectal cancer based on single-cell sequencing analysis and transcriptome analysis. Sci Rep (2026). https://doi.org/10.1038/s41598-025-34933-0
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DOI: https://doi.org/10.1038/s41598-025-34933-0
